DNA methylation is crucial for gene silencing, cell differentiation, and development. Although the mechanisms of DNA methylation have been well studied, those of DNA demethylation have been elusive. It has been reported that DNA demethylation of the Hes5 and GFAP promoters occurs during early neurogenesis and gliogenesis, respectively. Therefore, it is thought that DNA demethylation is an important event for neural and/or glial differentiation. Recent studies indicate that Tet family proteins (Tet1, Tet2 and Tet3) are able to catalyze the conversion of 5-methylcytosine of DNA to 5-hydroxymethylcytosine, suggesting a potential role for Tet proteins in DNA demethylation. We predicted that Tet family proteins could be involved in neural and/or glial differentiation. Here we identified Tet3 regulates not neural differentiation but glial differentiation in neuroblastoma Neuro2A cell line. Tet3 mRNA but not Tet1 and Tet2 were major transcripts in Neuro2A cells, In addition, Tet3 but not Tet1 nor Tet2 mRNA levels were increased more than 2.5-fold during retinoic acid (RA)-induced differentiation. Tet3 knockdown significantly impaired RA-induced glial marker gene GFAP, although Tet3 knockdown did not affect the expression level of neural differentiation marker genes such as NeuroD1 and NeuN. On the other hand, Tet3 overexpression also disturbed GFAP expression level. These results indicate that precise amount of Tet3 protein plays an important role for glial differentiation.